Multiple Myeloma (MM) is a malignancy that initially responds to standard of care agents. Unfortunately, however, all patients relapse with disease that is refractory to further treatment. We hypothesize that progression and maintenance of MM requires balanced CD44 signaling that controls homeostasis of intracellular Ca2+ pools, as we have shown that a first in class, in-house small molecule selective agonist of CD44 coined MTI-101 provokes toxic increases in the levels of intracellular Ca2+ that trigger MM cell necrosis. Further, we have identified key effectors downstream of CD44 that appear to control intracellular Ca2+ levels and to contribute to MTI-101-induced necrotic cell death, specifically an endoplasmic reticulum (ER) resident thiol oxidoreductase coined Ero1L and its target Stim1, an ER/plasma membrane (single-pass) calcium sensor. Importantly, we have also shown that: (i) MTI-101 has activity against both primary and relapsed multiple myeloma; (ii) Ero1L expression in MM patient samples correlates with sensitivity to MTI-101; and (iii) knockdown of Ero1L augments the sensitivity of MM to bortezomib-induced cell death. In Specific Aim 1 we will fully interrogate the status of the CD44-Ero1L-Stim1-Ca2+ circuit in primary myeloma specimens (MGUS, newly diagnosed and proteasome inhibitor refractory disease) and will assess effects of CD44 engagement by MTI-101 versus its natural ligand hyaluronic acid (HA) on intracellular Ca2+ levels and MM cell survival. We will also evaluate if levels of Ero1L and Stim1 in primary and relapsed MM correlate with sensitivity to MTI-101, with changes in intracellular Ca2+ pools following CD44 engagement, and with clinical resistance to bortezomib treatment. In Specific Aim 2 genetic and biochemical approaches will be used to assess the regulation and roles of Ero1L and Stim1 in the response to MTI-101 versus HA, and their roles in MTI-101 and bortezomib efficacy as therapeutics for myeloma in vivo. Finally, in Specific Aim 3 we will use mathematical models and validation studies to optimize single agent and combination therapy with MTI-101. We submit the proposed studies will establish the mechanism of action and efficacy of this exciting new agent that targets CD44-Ero1L- Stim1-Ca2+ circuit in both primary and relapsed multiple myeloma.